The ?-adrenergic receptor-adenyl-cyclase system of rat reticulocytes

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Abstract
Non-nucleated red blood cells from rats contain adenyl cyclase, the activity of which is predominantly localized in the reticulocytes. Basal enzyme activities in membrane preparations from reticulocyte-rich blood (pretreatment of rats with acetyl-phenylhydrazide: about 60% reticulocytes) are about 5 times higher than in preparations from reticulocyte-poor blood (untreated animals: 2–3% reticulocytes). The enzyme activities are stimulated 10-fold by sodium fluoride (10−2 M) and 6 to 8-fold by isoprenaline (10−4 M). Adenyl cyclase activities in membrane preparations from reticulocyte-rich and reticulocyte-poor blood can be ascribed to identical enzymes since identical apparent K m (ATP; 3×10−4 M), K a (isoprenaline; 3×10−6 M) and K i (propranolol vs. isoprenaline; 3×10−7 M) values were obtained in both preparations. Besides NaF, only phenylethanolamine derivatives with β-adrenergic receptor stimulant properties were effective as stimulators of adenyl cyclase activity. The affinities (apparent K a values) of the investigated compounds decreased in the order isoprenaline—hexoprenaline—fenoterol—salbutamol—adrenaline—terbutaline—noradrenaline—phenylephrine. For maximal intrinsic activity, the catechol structure was essential; the relative intrinsic activities of resorcinol derivatives did not exceed 0.6. The isoprenaline-stimulated adenyl cyclase activities in erythrocyte membrane preparations were competitively inhibited by β-adrenergic blocking drugs, the affinities (apparent K i values) decreasing in the order prindolol—penbutolol—propranolol—practolol. The dextrorotatory enantiomers of penbutolol and propranolol were 1/100 to 1/200 as active as the resp. levorotatory enantiomers. From experiments with α-adrenergic agonists (e.g. phenylephrine) and antagonists (e.g. phentolamine), it is concluded that α-adrenergic receptors do not interfere with the β-adrenergically-mediated cAMP formation in these particular membranes. A variety of hormones and drugs known to stimulate adenyl cyclase activities in various tissues, e.g. ACTH, glucagon, STH, erythropoietin, prostaglandin E 1 etc. did not affect adenyl cyclase activity in reticulocyte-rich erythrocyte membrane preparations. In contrast to adenyl cyclase activity, phosphodiesterase activities in erythrocyte membrane and cytoplasmic fractions were only twice as high in reticulocyterich as in reticulocyte-poor preparations. From the experiments described, it is obvious that the adenyl cyclase of the rat reticulocyte is subject to monovalent-hormonal, i.e. β-sympathomimetic stimulation. Moreover, the premature red blood cell provides a useful model for quantitative studies of the interaction of drugs with the β-adrenergic receptor.